四种短体线虫的形态和分子生物学鉴定

Morphological identification and molecular characteristics and phylogenetic relationship of four root lesion nematodes(Nematoda: Pratylenchidae)

采用形态学和分子生物学方法对来自荷兰的藏橐吾和铃兰、泰国的高山榕、缅甸的香蕉等种苗(球)分别鉴定出玻利维亚短体线虫Pratylenchus bolivianus、铃兰短体线虫P.convallariae、咖啡短体线虫P.coffeae和斯佩奇短体线虫P.speijeri。对这4种线虫的形态特征进行较为详细的描述后,认为头环、口针、侧区、生殖系统和尾形等形态特征是种类鉴定的重要依据;进一步利用引物28S-D2A/28S-D3Br扩增测序得到上述4种线虫的28S rRNA基因D2/D3区序列,序列分析发现玻利维亚短体线虫从欧洲到美洲的不同种群之间的遗传距离变异较小,仅为0~0.007;铃兰短体线虫同一种群不同个体之间的遗传变异较大,为0.006~0.029;咖啡短体线虫不同种群之间的平均遗传距离为0.013;斯佩奇短体线虫不同种群之间的平均遗传距离为0.010;咖啡短体线虫P.coffeae和斯佩奇短体线虫P.speijeri亲缘关系很近,二者种间平均遗传距离仅为0.026。本研究再次证明线虫28S rRNA基因D2/D3区基因序列可作为短体线虫种间鉴定的依据。

In our study,four species of root lesion nematodes including Pratylenchus bolivianus,P.convallariae,P.coffeae and P.speijeri were identified by morphology and molecular biology from Ligularia rumicifolia and Convallaria majalis in the Netherlands,Ficus altissima in Thailand,and Musa sp.in Myanmar,respectively.The morphological characteristics of these four nematode species were described in detail.Morphological characteristics,including head ring,stylet,lateral field,reproductive system and tail shape,etc.,are of great importance in terms of species classification.The sequences of the 28 S rRNA gene D2/D3 were obtained by using the primer pair 28 S-D2 A/28 S-D3 Br.The sequence analysis revealed that the genetic distance variation among the different populations of P.bolivianus from both Europe and America was small(0-0.007).By contrast,significant genetic variation of different individuals in the same population of P.convallariae was identified(0.006-0.029).The average genetic distance of different populations of P.coffeae was 0.013,while the average genetic distance between the different populations of P.speijeri was 0.010.In addition,the genetic relationship between P.coffeae and P.speijeri was close with each other,with the average genetic distance between them being only 0.026.Moreover,our study demonstrated that the D2/D3 gene sequence of the 28 S rRNA gene of nematode could be used as a basis for interspecific identification of root lesion nematodes.